The marginal band (MB) of nucleated erythrocytes is a curved microtubule system found just beneath the plasma membrane, relatively little studied but believed to play an important role in cell shape maintenance. Normally obscured by hemoglobin (Hb) in both living cells and TEM thin sections (due to Hb iron content), initial experiments have shown that it is stabilized and rendered easily visible by hemolysis in modified microtubule polymerization media containing EGTA and Triton X-100 (TXM). The "semi-lysed" erythrocyte thus produced consists of MB and nucleus, plus intra-MB material apparently holding the nucleus in native position. Both MB and intra-MB material are Ca ions-labile, and such experiments demonstrate that the semi-lysed cell is readily accessible to experimental media and microscopic investigation. The general objective of the proposed work is to exploit this system in studying the structure, composition, assembly/disassembly, and functional properties of the MB, and to test the intra-MB material for possible kinship to contractile microfilament systems. Experiments will be conducted on MB lability, MB isolation in structurally intact condition, on the possibly species-specific mirror-image twisting observed in TXM, on induction of contractile responses to ATP in the system, on MB function and differentiation during erythrocyte development and nuclear elimination, and on the correlation of results in vitro with experimentally induced shape changes in living erythrocytes by various agents including microtubule inhibitors and Ca ion ionophore. The work involves comparative observations on erythrocytes of a variety of species, and applies phase contrast light microscopy, transmission EM, scanning EM, and SDS-polyacrylamide gel electrophoresis to these problems. The data should be of value in understanding erythrocyte properties in general, blood platelets (which have MBs), and many other related systems.